The studies on the complementing fragment of proteins in this Section have led to the hypothesis that the interatomic interactions of the ordered structure of proteins would be globally coupled to generate extra energy, resulting in shift of the equilibrium of folding and unfolding in favor of folding. In application of this theory to the antigen-binding we speculate that not only the amino acid residues of the antibody which constitute the binding site but also the distant residues would play a role in providing extra force, through coupling, for the antigen-binding. If this is the case, elucidation of such global coupling would be important for understanding the antigen-binding. To investigate whether the global coupling is involved in the antigen-binding, we are preparing monoclonal antibody directed to yeast cytochrome c. The spleen cells of immunized BALB/c mice were fused with mouse myeloma (SP2/0 cell line) using polyethylene glycol. Three hybridomas (1-16-1, 9-29-20 and 39-14) hve been cloned, all producing IgMs. These IgMs were partially purified using Con A Sepharose and Sephacryl S300 or ammonium sulfate fractionation. All appear to have the antigenic determinants between residues 44 and 69 as determined using various fragments of yeast cytochrome c and enzyme-immuno assay. None reacted with horse or tuna cytochrome c or their apoproteins. However, all of them strongly react with yeast heme fragment (1-69)H as much as with native yeast cytochrome c but not with yeast heme fragment (1-43)H. Interestingly, antibody from clone (39-14) reacted with yeast as well as Candida krusei apocytochromes c and in the lesser extent with C. krusei native cytochrome c. The latest immunization with native yeast cytochrome c, glutaldehyde polymerized yeast cytochrome c and hemocyanci-coupled yeast cytochrome c, all have resulted in mice with the active TgG. Hydrodomas producing IgG have been obtained using one of these mice.